Localization of the site(s) of conversion of proglucagon to glucagon at the subcellular level. The recent discovery that hyperglucagonemia is inherent in all diabetics has stimulated new interest in learning more about alpha2 cell (glucagon producing) metabolism. It has become important to characterize modes of and factors controlling glucagon synthesis and release at an information level commensurate with and beyond that already available for insulin synthesis and release. Data from several studies including our own using anglerfish islet tissue indicates that glucagon, in a manner similar to insulin and most other peptide hormones, is synthesized via a larger precursor molecule. Results from preliminary experiments where isotope incorporation was followed by cell fractionation indicate that the microsome fraction is the biosynthetic site for proglucagon and proinsulin. Although much of the precursor to product conversion occurs after transfer of precursors to the secretory granule fraction, results from these experiments also indicate significant amounts of insulin and glucagon appear in the microsomal fraction with increasing incubation time. The goals of the proposed research are to attempt to specifically localize the site(s) of precursor conversion and thus the subcellular distribution of the enzyme(s) involved in precursor cleavage. Although the results obtained will not be directly applicable to treatment of diabetes, they should provide a better understanding of proteolytic processing of newly synthesized peptides and thus provide a basis for further research into the underlying defects in diabetic alpha cell metabolism.